Given the recent increase in cannabis use in the US, there is an urgent need to more clearly understand the effects of cannabinoids on central and peripheral signaling mechanisms. The main psychoactive component in cannabis, ?9-tetrahydrocannabinol (THC), has been shown to act directly on the cannabinoid receptors. These receptors are expressed in both central and peripheral cellular populations that have been shown to secrete extracellular vesicles, including the choroid plexus, neurons, glia, and adipocytes. The main goals of this proposal are to investigate the effects of THC on extracellular vesicle signaling in the brain and blood and to identify a panel of biomarkers related to THC use. To further validate the selectivity of the biomarker profile, RNA expression patterns will be compared in the presence of another drug commonly co-used with cannabis, nicotine. Thereafter, we will determine whether THC's effects on extracellular vesicle density and RNA cargo can be attributed to release from dopaminergic cells in the ventral tegmental area, a brain region shown to mediate the reinforcing and rewarding properties of drugs of abuse. By systematically examining changes in extracellular vesicle signaling with THC exposure and relating to drug co-use conditions, these studies seek to reveal important insights into central and peripheral extracellular signaling dynamics. Further, given that peripheral miRNAs have been proposed as biomarkers of disease state, and THC-containing cannabinoids are often used by patients, such as individuals with cancer, findings may further inform on the relevance of other proposed biomarker profiles for clinical applications. Finally, with this foundation, cannabinoid-associated extracellular signaling factors may also lead to the identification of extracellular mechanisms underlying drug dependence and thus potential novel targets for therapeutic development to treat dependence.